Recent studies identify the glucagon receptor (GR) as a critical contributor to hyperglycemia in type-2 diabetes. GR drives hepatic glucose production by initiating glycogenolysis in response to glucagon stimulation. Mutations in the GR gene impair endogenous glucagon feedback, leading to excessive glycogenolysis or gluconeogenesis and promoting type-2 diabetes onset. Glucagon resistance lowers blood glucose levels. These findings indicate tight regulation of the GR signaling pathway, and reducing hepatic glycogenolysis as a promising therapeutic strategy. Several regulatory components in this pathway remain unidentified. α-Arrestins are known to regulate membrane receptor internalization and membrane-protein homeostasis. This study investigates the role of α-arrestin protein ARRDC4 as a potential key modulator of GR trafficking and signaling. ARRDC4 expression is upregulated by high glucose and downregulated by insulin in hepatocytes. Our preliminary results from live cell imaging, using custom analysis tools, in hepatic (HepG2) cells indicates that ARRDC4 overexpression enhances GR surface expression and alters downstream endocytosis. This altered trafficking impacts downstream glucagon signaling and glycogenolysis, playing a key role in restoring glucose balance. In our approach, the hypothesis that ARRDC4 is a fundamental regulator of GR localization and signaling dynamics is further tested by a CRISPR ARRDC4 knockout HepG2 cell line generated to assess GR membrane expression and signaling upon glucagon stimulation using live cell imaging. Real-time downstream signaling activation is quantified based on cAMP activity using FRET biosensors. Findings were compared across ARRDC4 wild-type, knockout, and overexpressing HepG2 cells. Through this study, we identify a novel ARRDC4-GR regulatory axis, and a key player in regulating metabolism.
Sreenivas et al. (Sun,) studied this question.